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Hi,
I followed the steps to analyze MAESTER data as stated here https://github.com/petervangalen/MAESTER-2021
Once I merged the whole BAM file of the scRNASeq dataset (reads for all chromosomes) and the BAM file of the MAESTER dataset (only reads for chrM) I ran maegatk this way:
maegatk bcall -b HQ_CBs.csv -c 20 -o NUEVO_maegatk -mr 3 -i GSM5534703_K562-BT142.bam -n sGSM5534703_K562-BT142 -z -ub UB -bt CB -so
I also opened the merged BAM file from scRNASeq and MAESTER to check the coverage along the chrM. chrM is very well covered. Here is the IGV image.
I also include the content of the final directory. final.zip
I'm not sure whether the problems comes from maegatk or any script from the MAESTER pipeline so I also opened an issue in the MAESTER github petervangalen/MAESTER-2021#6
Any help would be much appreciated.
Regards,
Sheila
The text was updated successfully, but these errors were encountered:
Hi,
I followed the steps to analyze MAESTER data as stated here https://github.com/petervangalen/MAESTER-2021
Once I merged the whole BAM file of the scRNASeq dataset (reads for all chromosomes) and the BAM file of the MAESTER dataset (only reads for chrM) I ran maegatk this way:
maegatk bcall -b HQ_CBs.csv -c 20 -o NUEVO_maegatk -mr 3 -i GSM5534703_K562-BT142.bam -n sGSM5534703_K562-BT142 -z -ub UB -bt CB -so
Here is the resulting plot:
![maester](https://private-user-images.githubusercontent.com/32428329/256162983-9fa9acef-7b8e-4691-b9f9-ad49258db7ae.png?jwt=eyJhbGciOiJIUzI1NiIsInR5cCI6IkpXVCJ9.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.-hTOYGNotaqnflompw3Ncm99sVytMtToPEdKRsxipNQ)
I saw similar peaks in the supplementary information of the MAESTER paper (Figure 7a https://static-content.springer.com/esm/art%3A10.1038%2Fs41587-022-01210-8/MediaObjects/41587_2022_1210_MOESM1_ESM.pdf) indicating that those peaks belonged to the scRNASeq data so I must be missing something related to the processing of the MAESTER data but so far I have not found the problem.
Here is what my BAM files look like before merging both files to apply maegatk.
scRNASeq BAM (SRA identifier SRR15598773):
SRR15598773.lite.1.127471761 0 chr1 10019 1 91M * 0 0 TAACCCTAACCCTAACCCTAACCCTAACCCTAACCCTAACCCTAACCCTAACCCTAACCCTAACCCTAACCCTAACCCTAACCCAAACCCA ??????????????????????????????????????????????????????????????????????????????????????????? NH:i:4 HI:i:1 AS:i:87 nM:i:1 RG:Z:scRNASeq:0:1:unknow_flowcell:0 RE:A:I xf:i:0 CR:Z:ATCTTCATCCATCAGA CY:Z:???????????????? CB:Z:ATCTTCATCCATCAGA-1 UR:Z:TTTCTCTTAGTG UY:Z:???????????? UB:Z:TTTCTCTTAGTG
MAESTER BAM (SRA identifier SRR15598774):
SRR15598774_6928992 16 chrM 1 255 59S181M * 0 0 CTGACGGGCCATCACGCCCACACCGCCCCCACGTTCCCCTGAAATCAGACCTCCCGAGGGATCACAGGTCTATCACCCTATTAACCCCTCACGGGAGCTCTCCATGCATGTGGTATTTTCGTCTGGGGGGTGTGCACGCGATAGCATTGCGAGACGCTGGAGCCGGAGCACCCTATGTCGCAGTATCTGTCTTTGATTCCTGCCTCATCCTATTATTTATCGCACCTACGTTCAATATTA ,,,,,F:FF:,FF,,,F:,F,:,,F:::,F,FFF,,::FF::,,,,,F,F,FF,FFF,,F,FF:F,F,,:,FFF:FFF,,F,,,FF,FF:,FF:F:FF::FFF,,FF:F,FFF,F,FFFFFFFFFFFFFF:FFF:,FFFFF,:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:,FFFFFFFFFFFFFFFFFF,FF,FFF:FFFFFFFFFFF,F,:,F,FFFF:FFFFFF,FFF:FFF:F NH:i:1 HI:i:1 AS:i:173 nM:i:3 CB:Z:TGGAGGATCTTGTTAC-1 UB:Z:CACTTATTGTTA
I also opened the merged BAM file from scRNASeq and MAESTER to check the coverage along the chrM. chrM is very well covered. Here is the IGV image.
![IGV_MAESTER](https://private-user-images.githubusercontent.com/32428329/256197091-481bd4c5-8f87-4bd7-8e9e-f090f9d46aa7.png?jwt=eyJhbGciOiJIUzI1NiIsInR5cCI6IkpXVCJ9.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.vqOlMMevZmUM-ssCXSfw7jBc-09VzZu5tSwsOnZL37k)
I also include the content of the final directory.
final.zip
I'm not sure whether the problems comes from maegatk or any script from the MAESTER pipeline so I also opened an issue in the MAESTER github petervangalen/MAESTER-2021#6
Any help would be much appreciated.
Regards,
Sheila
The text was updated successfully, but these errors were encountered: